Abraxis Method 546 Guide

Transcription

1 Abraxis Method 546 Guide

2 Analysis Batch Controls Laboratory Reagent Blank Description: Control that is used to determine if any interferences are introduced during analysis. It is an aliquot of reagent water that is lysed and filtered using the same procedure used for sample processing Preparation Guidelines If drinking water samples are being analyzed with the batch, use DI/Distilled water treated appropriately with sodium thiosulfate If only testing raw water, do not treat DI/Distilled water with sodium thiosulfate Location on plate: LRB will be analyzed twice, once immediately after the standards and once at the end of the plate. Acceptable range: Must be 0.15 ppb (less than one half the MRL)

3 Analysis Batch Controls Laboratory Reagent Blank: Preparation Measure 10 ml of DI H 2 O into two 20 ml vials Three Freeze/Thaw Cycles Filter each separately with glass fiber filters

4 Analysis Batch Controls Low-CV Description: Control that is at a concentration that is less than or equal to the MRL (0.3 ppb) which is used to confirm the accuracy of the standard curve near the MRL. Preparation Guidelines Do not freeze/thaw or filter the Low-CV Preparation: May be obtained by Abraxis in the Method 546 Accessory Pack Location on plate: Low-CV will be analyzed once, immediately after the first LRB Acceptable range: Must be 0.15 or 0.45 ppb

5 Analysis Batch Controls Low-CV: Preparation Measure 10 ml of DI H 2 O into a single 20 ml vial Add 60 ul of the Abraxis 50 ppb standard into the vial Vortex or invert repeatedly to mix DO NOT FREEZE/THAW OR FILTER THE LOW-CV

6 Analysis Batch Controls Laboratory Fortified Blank Description: An aliquot of reagent water to which a known concentration (0.6 ppb) is added to verify method performance in the absence of sample matrix. Preparation Guidelines If finished drinking water samples are included in the batch, properly quenched DI H 2 O must be used to prepare the LFB. If only analyzing raw drinking water, do not quench the DI H 2 O. Location on plate: Immediately after the Low-CV and again at the end of the plate immediately after the last sample. Acceptable range: Must be 0.36 or 0.84 ppb

7 Analysis Batch Controls Laboratory Fortified Blank: Preparation Measure 10 ml of DI H 2 O into two separate 20 ml vials Pipette 120 ul of the Abraxis 50 ppb standard into each sample vial Three Freeze/Thaw Cycles Filter each separately with glass fiber filters

8 Analysis Batch Controls Laboratory Fortified Sample Matrix and LFSM Duplicate Description: Aliquot of a new or previously analyzed sample to which a known concentration of microcystins is added. It is used to determine if the sample matrix contributes bias to the results. Preparation guidelines: If analyzing only drinking water, a quenched sample must be used to prepare the LFSM and LFSMD. If only analyzing raw water, a non-quenched sample must be used to prepare the LFSM and LFSMD. If analyzing both raw and finished drinking water, a representative set of LFSM and LFSMDs must accompany the samples. If more than 20 samples are to be analyzed of either type, a second representative set of LFSM and LFSMD must be analyzed. Location on plate: Immediately after the first LFB. If analyzing more than one set of LFSM and LFSMD, the second set will follow half of the samples. Acceptable range: Calculate the mean percent recovery (%R) for each LFSM and LFSMD set using the equation: Where, A = mean measured concentration of the LFSM and LFSMD, B = measured concentration in the unfortified sample C = fortification concentration (0.5 ppb for drinking water, 1.0 ppb for ambient water). Subtract the unfortified sample recovery (B) from the mean fortified spiked recovery (A) even if the concentration in the unfortified sample is less than the MRL (0.3 ppb). Calculate the relative percent difference (RPD) between the LFSM and LFSMD using the following equation: The %R for the LFSM and LFSMD must be 60% and 140%. The RPD for the LFSM and LFSMD should be 40%.

9 Analysis Batch Controls LFSM and LFSMD Acceptability Criteria Calculate the mean percent recovery (%R) for each LFSM and LFSMD set using the equation: Where, A = mean measured concentration of the LFSM and LFSMD, B = measured concentration in the unfortified sample C = fortification concentration (0.5 ppb for drinking water, 1.0 ppb for ambient water). Subtract the unfortified sample recovery (B) from the mean fortified spiked recovery (A) even if the concentration in the unfortified sample is less than the MRL (0.3 ppb). Calculate the relative percent difference (RPD) between the LFSM and LFSMD using the following equation: The %R for the LFSM and LFSMD must be 60% and 140%. The RPD for the LFSM and LFSMD should be 40%.

10 Analysis Batch Controls LFSM/LFSMD: Preparation (Drinking Water) Select a new or previously quenched sample, invert several times to mix and measure 10 ml into two separate 20 ml sample vials Pipette 100 ul of the Abraxis 50 ppb standard into each sample vial, and vortex or invert repeatedly to mix Three Freeze/Thaw Cycles Filter each separately with glass fiber filters

11 Analysis Batch Controls LFSM/LFSMD: Preparation (Raw Water) Select a new or previously analyzed sample, invert several times to mix and measure 10 ml into two separate 20 ml sample vials Pipette 200 ul of the Abraxis 50 ppb standard into each sample vial, and vortex or invert repeatedly to mix Three Freeze/Thaw Cycles Filter each separately with glass fiber filters

12 Analysis Batch Sample Plate Map A B C D E F G H Std. 0 Std. 4 LFB ppb 2.0 ppb 0.60 ppb Std. 0 Std. 4 LFB ppb 2.0 ppb 0.60 ppb Std. 1 Std ppb 5.0 ppb Std. 1 Std ppb 5.0 ppb S2 S6 S10 S14 LFSMD S20 S24 S28 S32 S2 S6 S10 S14 LFSMD S20 S24 S28 S32 LFSM S3 S7 S11 S15 S17 S21 S25 S29 S33 LFSM S3 S7 S11 S15 S17 S21 S25 S29 S33 Std. 2 LRB 1 LFB ppb < 0.15 ppb LFSMD S4 S8 S12 S16 S18 S22 S26 S ppb Std. 2 LRB 1 LFB ppb < 0.15 ppb LFSMD S4 S8 S12 S16 S18 S22 S26 S ppb Std. 3 Low-CV LRB ppb 0.30 ppb S1 S5 S9 S13 LFSM S19 S23 S27 S31 < 0.15 ppb Std. 3 Low-CV LRB ppb 0.30 ppb S1 S5 S9 S13 LFSM S19 S23 S27 S32 < 0.15 ppb NOTES: Additional sets of LFSM/LFSMD may be required depending upon the presence of both ambient and drinking water or if greater than 20 samples of either type are analyzed. A 546 Quality Control Sample (546 QCS) must be analyzed with each new lot of calibrators. Place it after the first set of LFSM/LFSMD at the beginning of the plate. In this example, replace both S1 replicates with the Method 546 QCS. The plate map above shows the analysis of an entire microtiter plate. As the number of samples on a plate increases, the potential for bias due to unequal incubation periods, otherwise known as drift, increases. To avoid the potential for drift, ensure that the addition of antibody, conjugate, color and stop are completed within two minutes. If any of these steps cannot be completed in less than two minutes, reduce the number of samples being analyzed on a single plate accordingly. To increase the speed at which the addition of these solutions are performed, a repeater pipette, 8 channel pipette or an automated analyzer (CAAS) can be used to load these reagents.

13 Initial Demonstration of Capability Controls Acceptable Sample Background Preparation guidelines: Use a portion of the remaining DI H 2 O treated with sodium thiosulfate prepared for the demonstration of precision and accuracy to prepare Acceptable Sample Background controls Location on plate: scattered throughout plate, first one after the Method 546 QCS Acceptability criteria: The result for each of the five LRBs must be less than one-half the concentration of the MRL, or 0.15 ppb.

14 Initial Demonstration of Capability Controls Acceptable Sample Background: Preparation Aliquot 5 ml of quenched DI H 2 O into five separate sample 20 ml sample vials Three Freeze/Thaw Cycles Filter each separately with glass fiber filters

15 Initial Demonstration of Capability Controls Precision and Accuracy Location on plate: Scattered throughout plate after the first Acceptable Sample Background control Acceptability Criteria: The % Relative Standard Deviation (%RSD) between the seven sets of well duplicates must be 15%. The mean recovery for the seven replicates must be 70% and 130% of the target value, or 0.35 and 0.65 ppb. A macro for the calculation of recoveries and %RSD can be obtained from Abraxis upon request.

16 Initial Demonstration of Capability Controls Precision and Accuracy: Preparation Measure 200 ml in an appropriate glass container and add two 10 mg tablets of sodium thiosulfate, mix well by inversion to dissolve Aliquot 10 ml of the above solution into seven separate 20 ml vials, along with 100 ul of the Abraxis 50 ppb standard. Mix well by inversion or vortex. Complete 3 Freeze/Thaw cycles, and use a glass fiber filter to filter each one in a separate vial

17 Initial Demonstration of Capability Controls IDC MRL Preparation guidelines: Use a portion of the remaining DI H 2 O treated with sodium thiosulfate prepared for the demonstration of precision and accuracy to prepare IDC MRLs Location: scattered throughout plate, first one after the first Acceptable Sample Background control Acceptability criteria: The Upper PIR Limit must be 150% and the Lower PIR Limit must be 50%. A macro for the calculation of recoveries can be obtained from Abraxis upon request.

18 Initial Demonstration of Capability Controls IDC MRL Aliquot 10 ml of treated DI H 2 O into seven separate 20 ml vials Pipette 60 ul of the Abraxis 50 ppb standard into each sample vial, and vortex or invert repeatedly to mix Three Freeze/Thaw Cycles Filter each separately with glass fiber filters

19 Method 546 QCS Analyze a Method 546 QCS, which is contained within the Abraxis Method 546 Accessory Kit, each time an IDC is attempted or when a new lot of calibration standards is used. Analyze the Method 546 QCS provided in the Abraxis 546 Accessory Pack with the IDC/Analysis Batch. No additional preparation is necessary. Do not use the QCS contained in the base Microcystins kit (PNs and OH). Do not add sodium thiosulfate, lyse or filter the 546 QCS

20 Initial Demonstration of Capability Sample Plate Map A Std. 0 Std QCS ASB LRB2 P&A3 IDC MRL4 P&A6 LFB 2 B Std. 0 Std QCS ASB LRB2 P&A3 IDC MRL4 P&A6 LFB 2 C Std. 1 Std. 5 ASB LRB1 P&A2 IDC MRL3 ASB LRB5 IDC MRL6 D Std. 1 Std. 5 ASB LRB1 P&A2 IDC MRL3 ASB LRB5 IDC MRL6 E Std. 2 Low-CV P&A1 IDC MRL2 ASB LRB4 P&A5 P&A7 F Std. 2 Low-CV P&A1 IDC MRL2 ASB LRB4 P&A5 P&A7 G Std. 3 LFB 1 IDC MRL1 ASB LRB3 P&A4 IDC MRL5 IDC MRL7 H Std. 3 LFB 1 IDC MRL1 ASB LRB3 P&A4 IDC MRL5 IDC MRL7 Key: ASB = Acceptable Sample Background LRBs, target value 0.15 ppb P&A = Precision and Accuracy Samples, target value 0.50 ppb IDC MRL = Initial Demonstration of Capability Minimum Reporting Level Sample, target value 0.30 ppb

21 Questions or Comments? Zak Hutchinson / Technical Support Specialist zhutchinson@abraxiskits.com Abraxis, Inc Office: (215) / Fax: (215) Railroad Drive Warminster, PA